Lee Sau Har, Jaganath Indu Bala, Manikam Rishya, Sekaran Shamala Devi
Department of Medical Microbiology, Faculty of Medicine, Universiti Malaya, Kuala Lumpur, Malaysia.
BMC Complement Altern Med. 2013 Oct 20;13:271. doi: 10.1186/1472-6882-13-271.
Lung cancer constitutes one of the malignancies with the greatest incidence and mortality rates with 1.6 million new cases and 1.4 million deaths each year. Prognosis remains poor due to deleterious development of multidrug resistance resulting in less than 15% lung cancer patients reaching five years survival. We have previously shown that Phyllanthus induced apoptosis in conjunction with its antimetastastic action. In the current study, we aimed to determine the signaling pathways utilized by Phyllanthus to exert its antimetastatic activities.
Cancer 10-pathway reporter array was performed to screen the pathways affected by Phyllanthus in lung carcinoma cell line (A549) to exert its antimetastatic effects. Results from this array were then confirmed with western blotting, cell cycle analysis, zymography technique, and cell based ELISA assay for human total iNOS. Two-dimensional gel electrophoresis was subsequently carried out to study the differential protein expressions in A549 after treatment with Phyllanthus.
Phyllanthus was observed to cause antimetastatic activities by inhibiting ERK1/2 pathway via suppression of Raf protein. Inhibition of this pathway resulted in the suppression of MMP2, MMP7, and MMP9 expression to stop A549 metastasis. Phyllanthus also inhibits hypoxia pathway via inhibition of HIF-1α that led to reduced VEGF and iNOS expressions. Proteomic analysis revealed a number of proteins downregulated by Phyllanthus that were involved in metastatic processes, including invasion and mobility proteins (cytoskeletal proteins), transcriptional proteins (proliferating cell nuclear antigen; zinc finger protein), antiapoptotic protein (Bcl2) and various glycolytic enzymes. Among the four Phyllanthus species tested, P. urinaria showed the greatest antimetastatic activity.
Phyllanthus inhibits A549 metastasis by suppressing ERK1/2 and hypoxia pathways that led to suppression of various critical proteins for A549 invasion and migration.
肺癌是发病率和死亡率最高的恶性肿瘤之一，每年有160万新发病例和140万人死亡。由于多药耐药性的有害发展，预后仍然很差，导致不到15%的肺癌患者存活五年。我们之前已经表明，叶下珠具有诱导细胞凋亡和抗转移作用。在本研究中，我们旨在确定叶下珠发挥其抗转移活性所利用的信号通路。
进行癌症10通路报告基因阵列，以筛选叶下珠在肺癌细胞系（A549）中发挥其抗转移作用所影响的通路。然后通过蛋白质印迹、细胞周期分析、酶谱技术和基于细胞的人总诱导型一氧化氮合酶酶联免疫吸附测定法对该阵列的结果进行确认。随后进行二维凝胶电泳，以研究叶下珠处理后A549细胞中的差异蛋白质表达。
观察到叶下珠通过抑制Raf蛋白来抑制ERK1/2通路，从而发挥抗转移活性。抑制该通路导致MMP2、MMP7和MMP9表达受到抑制，从而阻止A549细胞转移。叶下珠还通过抑制HIF-1α来抑制缺氧通路，导致VEGF和诱导型一氧化氮合酶表达降低。蛋白质组学分析揭示了一些被叶下珠下调的蛋白质，这些蛋白质参与转移过程，包括侵袭和迁移蛋白（细胞骨架蛋白）、转录蛋白（增殖细胞核抗原；锌指蛋白）、抗凋亡蛋白（Bcl2）和各种糖酵解酶。在所测试的四种叶下珠物种中，余甘子显示出最大的抗转移活性。
叶下珠通过抑制ERK1/2和缺氧通路来抑制A549细胞转移，这导致抑制了A549细胞侵袭和迁移的各种关键蛋白。
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